Characterization of Caulobacters isolated from wastewater treatment systems and assay development for their enumeration

Caulobacters are gram-negative bacteria that have a biphasic life cycle consisting of a swarmer and a stalked stage. As a result they have elicited interest as a simple developmental model. Less attention has focussed on their role in the environment, although they have been found in almost every aquatic environment as well as in many soils. Caulobacters are often described as oligotrophic bacteria because of their prevalence in pristine waters but have now been isolated from the relatively nutrient-rich wastewater environment. In order to learn more about this population some basic characterization was carried out and an assay system to determine their prevalence in sewage plants was designed. Most of the organisms isolated from sewage treatment facilities had similar gross morphological features, but differed in holdfast composition, total protein profile, antibiotic resistance and restriction fragment length polymorphism, thereby indicating a greater diversity than originally assumed. Most of the organisms hybridized with flagellin and surface array genes that had previously been cloned, and only one of 155 non-Caulobacter sewage isolates hybridized with the flagellin gene probe; consequently these were used in a DNA-based enumeration strategy. DNA was isolated directly from sewage and probed with the flagellin and the surface array gene probes. The signals obtained were compared to standards made up of pooled Caulobacter DNA from the sewage isolates and non-Caulobacter DNA from organisms also present in sewage. Using this assay Caulobacters could only be detected above the 1% level, which was higher than their proportion in the wastewater environment. It appears that this approach will not be useful in monitoring Caulobacters in treatment plants unless a more highly conserved or higher copy number probe is found.Science, Faculty ofMicrobiology and Immunology, Department ofGraduat